Ras is a key molecule in the transduction of signals, from the cell surface to the nucleus, which regulate essential cellular processes such as proliferation, differentiation, gene expression, and death. Activating point mutations affect the normal transduction of signals that result in the development of many disorders and are present in 30 percent of all tumors. Understanding the role each Ras isoform plays in steady state signaling pathways, such as those leading to apoptosis, will help explain how perturbations in these pathways leads to disease development. Preliminary data suggest that c-K-Ras knockout murine fibroblasts are more resistant to the apoptotic agent, TNF-a, than K+/+ cells. The investigation outlined in this proposal will test the hypothesis that c-K-Ras is involved in apoptotic signaling pathways. This hypothesis will be tested through the execution of two specific aims. Specific Aim 1: Determine the universal nature of pro- versus anti-apoptotic signals generated by c-K-ras gene products, c-K-Ras 4A (KA) and 4B (KB), and the involvement of each isoform in the apoptotic pathways. The sensitivity of K-/- vs. K+/+ murine fibroblasts to apoptotic inducing agents will be determined. The contribution of KA and/or KB to these signals will be evaluated. Specific Aim 2: Determine the signaling pathways downstream of c-K-Ras isoforms that impinge on known regulators of apoptotic pathways. We will map the activation of typical Ras effector pathways and determine the contribution of each pathway to the apoptotic response using c-K-Ras mutants that activate discrete effectors.